Previously, complete sequences for the mRNAs encoding the RSV G and F glycoproteins were determined from cDNA clones. Here we constructed vaccinia virus recombinants (vaccinia-F and vaccinia-G) that express the individual RSV glycoproteins. Vaccinia-F and vaccinia-G were administered to cotton rats to evaluate the relative contributions of the individual glycoproteins to host immunity. Both recombinants induced high levels of RSV serum antibodies, measured both by ELISA and by RSV neutralization plaque reduction. Upon subsequent intranasal (IN) challenge with RSV, vaccinia-G provided substantial protection in the lungs and no protection in the nose, and vaccinia-F provided complete protection in the lungs and partial protection in the nose. Importantly, the serum neutralizing antibody titers induced by vaccinia-F were slightly higher than that induced by RSV infection of the respiratory tract, and 6-fold higher than obtained with vaccinia-G. Thus, the F glycoprotein appears to be more effective in stimulating host immunity. These results show that the vaccinia virus recombinants should be evaluated further as potential methods of immunoprophylaxis. Also, the cotton rat was found to be a sensitive assay for testing the safety of vaccinia virus recombinants. The F and G cDNAs have also been expressed in tissue culture using recombinant SV40 viruses. In conjunction with site-directed mutagenesis, these studies will define amino acid sequences that are important in protein function.